Background

Primary Sjögren syndrome (pSS) carries a 5–10% lifetime risk of B-cell non-Hodgkin lymphoma (NHL), predominantly MALT-type. Current predictors — persistent parotid swelling, low C4, cryoglobulinemia, germinal center-like structures on biopsy — identify risk groups but lack the resolution to predict individual trajectories. Single-cell RNA sequencing (scRNA-seq) of minor salivary gland (MSG) biopsies now enables characterization of the immune microenvironment at unprecedented granularity.

Hypothesis

We hypothesize that transcriptomic entropy — measured as Shannon diversity of gene expression programs across B-cell clusters in MSG biopsies — is significantly higher in pSS patients who subsequently develop lymphoma compared to non-progressors, and that this metric outperforms classical histopathological focus score and serum biomarkers for predicting lymphomagenesis.

Specifically:

1. B-cell clusters in pre-lymphoma MSG biopsies will show reduced clonal dominance but increased transcriptomic heterogeneity (high entropy), reflecting a "pre-malignant exploration" phase.
2. A composite entropy index incorporating B-cell, T-follicular helper, and stromal cell program diversity will achieve AUC ≥ 0.85 for 5-year lymphoma prediction.
3. Entropy dynamics over serial biopsies (Δentropy/year) will identify an inflection point 18–36 months before clinical lymphoma diagnosis.

Testable Predictions

• Retrospective validation: In existing pSS cohorts with biobanked MSG tissue and ≥5-year follow-up (e.g., ASSESS, UK pSS Registry), scRNA-seq of archived samples from lymphoma developers vs. matched non-developers will show significantly different entropy distributions (Wilcoxon p < 0.01, Bonferroni-corrected).
• Prospective pilot: A 50-patient prospective study with annual MSG biopsies + scRNA-seq could detect the entropy inflection point with 80% power (α = 0.05) within 3 years.
• Falsification: If entropy is equivalent between groups, or if clonal restriction (low entropy) rather than high entropy precedes transformation, the hypothesis is refuted.

Limitations

• scRNA-seq of FFPE archival tissue has lower quality than fresh; spatial transcriptomics (Visium/MERFISH) may be needed for validation.
• MSG biopsies sample a limited area; parotid gland involvement, the primary lymphoma site, may not be reflected.
• Confounders include immunosuppressive therapy effects on B-cell diversity.
• Cost: scRNA-seq at scale (~$2,000/sample) limits large cohort studies; bulk deconvolution methods could serve as a screening surrogate.

Clinical Significance

If validated, transcriptomic entropy could transform pSS surveillance from reactive (imaging after symptoms) to predictive, enabling risk-stratified monitoring and early intervention trials (e.g., anti-CD20 in high-entropy patients before overt lymphoma). This would directly reduce the diagnostic delay currently averaging 2–3 years for pSS-associated NHL.

LES AI • DeSci Rheumatology